Calorie restriction (CR) has an antitumorigenic effect against triple-negative breast cancer (TNBC). A key mitogenic pathway modulated by CR involves insulin-like growth factor (IGF)-1, which plays a central role in local and systemic growth and cell survival by activating the PI3K/AKT/mTOR pathway downstream of the IGF-1 receptor (IGF-1R). Although multiple miRNAs that target components of the IGF-1 signaling pathway have been identified, the impact of CR on IGF-1-related miRNA expression has not been addressed. We hypothesize that CR-induced decreases in TNBC development and progression involve miRNA's regulating IGF-1 signaling, we used C3(1)/SV40 T-antigen (C3-TAg) mice (a TNBC model), a C3-Tag progression series of cell lines (M28, M6, and M6C), and human TNBC cell line (MDA-MB-231). The expression of miRNAs previously reported to target components of the IGF-1 and/or mTOR pathways was evaluated by real-time PCR. Hormone levels were measured using the mouse Luminex Screening Assay. To mimic CR in vitro, and to test the effects of miRNA manipulation, the C3-Tag series of TNBC cell lines were treated (separately and in combination) with serum restriction, BMS754807 (inhibitor of IGF1-R), and various levels of recombinant IGF-1 as well as manipulated levels of miRNA's associated with IGF-1/mTOR signaling. We also conducted in silico analysis using Metabric, TCGA, TargetScan, and miRTarBase data sets to identify genes and pathways associated with specific miRNAs expression related to the PI3K/Akt/mTOR pathway. Our results confirm that CR maintains lower body weight, reduces circulating levels of IGF-1, and slows spontaneous TNBC development and progression in mice. CR also significantly increased the expression of miR-15b and miR-486. In vitro studies showed miR-15b and miR-486 expression decreased with increasing tumorigenicity of the C3-tag series of TNBC progression. Treatment of mouse and human TNBC cells with low IGF-1 induced miR-15b expression and inhibited proliferation. Moreover, miR-15b overexpression inhibited cell proliferation and decreased IGF-1R expression (mRNA and protein). Consistent with these results, the in silico analysis shows that the amplification of miR-15b correlates with downregulation of pathways involved in tumor growth and IGF1-R signaling. Together these findings suggest that reduced circulating IGF-1 levels in response to CR leads to the upregulation of miR-15b, which correspondingly targets and downregulates IGF-1R, both at the mRNA and protein levels. This combination of reduced IGF-1 ligand and miR-15b-induced IGF-1R downregulation contributes to the potent anticancer effects of CR and reveals potential targets for pharmacologically mimicking those effects. This research is supported by R35CA197627.
Citation Format: Ximena Minerva Bustamante-Marin, Kaylyn L. Devlin, Om Dave, Jenna L. Merlino, Shannon B. McDonell, Michael F. Coleman, Stephen D. Hursting. Anticancer effects of calorie restriction in a murine C3-TAg model of triple-negative breast cancer: the role of miR-15b [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2357.